bacterial enumeration gloria phuong le microbiology lab dr. fran norflus
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Bacterial Enumeration
Gloria Phuong LeMicrobiology LabDr. Fran Norflus
Introduction
Bacteria divide via binary fission but not all daughter cells are viable
Bacterial enumeration is the measurement of bacterial cells per unit of sample (volume or mass)
Counting of bacterial cells is important in: Microbial ecology Epidemiology
Methods
Total count vs. viable count: all cells vs. viable cells capable of producing progeny
There are 2 ways of counting bacterial cells: direct and indirect Direct methods of enumeration involve counting
actual cells or colonies Indirect methods involve estimating the number of
cells based on cell mass, scattering of light through a culture (spectroscopy), or through statistical method called most probable number (MPN)
Methods
Method Enumeration Description
Indirect and viable MPN Statistical estimation based on growth patterns
Direct and viable Standard plate count Serial dilution of original sample and look for CFU
Indirect and total Spectroscopy Measure the transmittance of sample
Direct and total Staining and microscopy
Stain the cells with fluorescent dyes to make them visible in raw samples
Microbes are everywhere
Foods can cause hazards to a person’s health if They possess a large number of bacteria that can cause
an infection by mere ingestion of the food Contains microbial products of metabolism that can cause
intoxication of the body
Food-handling techniques are important to prevent food poisoning We will examine the presence of microbes in chopped beef
and chicken samples Normal and abnormal handling
Direct and viable method would be used
Procedures
Dilute the original sample (serial dilution)
Then use 0.1 ml of each dilution to perform the spread plate method (p. 56-59)
The plates will be incubated at 37oC for 48 hours
Spread Plate Method
Direct and viable count
After 48 hours, there will be colonies growing on the plates
We will perform standard plate count (direct method to determine viable cells)
Each colony originated from one mother cell
We will count colony-forming-unit (CFU)
Record in your notes either TNTC: too numerous too count The exact CFU: 30-300
How to perform calculations?
First, we need to determine the concentrations of cells from countable plates
Then, we need to determine the dilution factor at each dilution (Df)
Next, we need to determine the total dilution (TD) TD=Df1xDf2xDf3…
Finally, to determine the concentration of cells in the original sample, we divide the number of CFU in the sample to the TD of that sample